The SLC4 family of membrane transporters mediate Cl--HCO3- and Na+-driven Cl--HCO3- exchange, and Na+- HCO3- cotransport (electrogenic, electroneutral). The importance of these transporters in mammalian cell biology is highlighted by the phenotypic abnormalities resulting from spontaneous SLC4 mutations in humans and targeted deletions in mice. Abnormalities in either the function and/or plasma membrane expression of certain members of the SLC4 family are the cause of various genetic diseases in man. Of the known SLC4 genes, SLC4A4 (NBCe1) and SLC4A1 (AE1) play key roles in mediating renal bicarbonate absorption, and the maintenance of systemic and intracellular acid-base balance. Mutations in NBCe1 impair both proximal tubule and extrarenal acid-base transport. In preliminary experiments, we have begun to examine the structural basis for the functional properties of NBCe1 and have made several novel discoveries that form the background for this proposal. Preliminary studies of the anion binding machinery of NBCe1 have led to the novel finding that it functions as both an electrogenic Na+-CO32- cotransporter and an electroneutral Na+-NO3- cotransporter. Mutations in NBCe1 that cause renal tubular acidosis also inhibit NO3- transport. Based on these new findings, NBCe1 must now be considered to be a bi-functional transporter that mediates carbonate and nitrate transport in the proximal tubule, and potentially in extrarenal tissues where it is expressed. Given our findings that NBCe1 transports Na+-CO32- and Na+-NO3-, the role of the kidney and the proximal tubule in inter-organ NO3- cycling and whole body NO3- balance becomes particularly relevant both from a physiologic and clinical standpoint. The proposal will characterize the architectural basis for the function of NBCe1 as both an electrogenic Na+-CO32- cotransporter and an electroneutral Na+-NO3- cotransporter, the perturbation of these functional properties in disease, and the role of NBCe1 and the proximal tubule in NO3- transport.